Bi-functional co-polymer use for ophthalmic and other topical and local applications

ABSTRACT

The invention contemplates a copolymer which is a graft or block copolymer useful to change wettability and surface characteristics of biological surfaces. Methods for use of these formulations and coatings to change wettability and sterically stabilize, and lubricate biological surfaces in a subject, for example, in the treatment of dry eye syndrome, and to prevent adherence of unwanted proteins, for example in the treatment of contact lens intolerance, are provided.

CROSS-REFERENCE TO RELATED APPLICATION

This application is a continuation of U.S. Ser. No. 14/328,484, filedJul. 10, 2014, which is a continuation of U.S. Ser. No. 12/708,329,filed Feb. 18, 2010, which claims the benefit of Provisional ApplicationNo. 61/153,416, filed on Feb. 18, 2009, the entire contents of each ofwhich are incorporated herein by reference in their entirety.

FIELD OF THE INVENTION

The present invention provides topical and/or local deliverypharmaceutical formulations and coatings for extraocular devices, drugparticles, or drug containing particles comprising a bi-functionalpolymer or co-polymer and methods for their use in modifying surfacecharacteristics and/or changing the wettability of biological surfacesin a subject, or on the aforementioned devices. Formulations of thepresent invention are particularly useful in various applicationsincluding hydration, in particular treatment of dry eye syndrome andtreatment of ophthalmic discomfort prevention of infection and/oradhesion of bacteria, viruses, proteins, toxins, and antigens anddelivery of active pharmaceutical ingredients to biological surfaces.Formulations of the present invention can be used to alter thetribological properties of biological surfaces. Formulations of thepresent invention are also useful when combined with a host of topicallydelivered therapeutic agents. The formulations are also useful forcoating extraocular devices, particularly contact lenses, to improvetheir biological compatibility.

BACKGROUND OF THE INVENTION

Poly(L-Lysine)-graft-poly(ethylene glycol) (PLL-g-PEG) is a watersoluble copolymer consisting of a poly(L-Lysine) backbone andpoly(ethylene glycol) side chains (Sawhney et al. Biomaterials 199213:863-870). The PLL chain, which carries multiple positive charges,spontaneously adsorbs onto negatively charged surfaces while PEG is apolynonion which serves as a non-binding domain. This adsorption isstrong and occurs rapidly, and renders surfaces protein and cellresistant. Furthermore, PLL-g-PEG has been shown to improve thebiocompatibility of materials. (Sawhney et al. Biomaterials 199213:863-870)

Various applications for the PLL-g-PEG graft co-polymer, blockco-polymers, and dendrimers with similar binding characteristics havebeen described.

SUMMARY OF THE INVENTION

In one aspect, the invention provides a method for increasing thewettability of a biological surface or treating biological surfacerelated discomfort in a subject, involving contacting the surface with acomposition containing a copolymer having a positively charged, orhydrophobic or covalent bonding moiety and a hydrophilic moiety, wherethe contacting is effected in an amount and for a duration so as toincrease the wettability or treat discomfort of the biological surface.

In one aspect, the invention provides a composition containing acopolymer having a positively charged, or hydrophobic, or covalentbonding moiety and a hydrophilic moiety and a pharmaceuticallyacceptable carrier.

In one aspect, the invention provides a pharmaceutical formulationcontaining an effective amount of a composition containing a copolymerhaving a positively charged, or hydrophobic, or covalent bonding moietyand a hydrophilic moiety and a pharmaceutically acceptable carrier.

In another aspect, the invention provides a method of storing a contactlens for at least 1 hour, involving providing the contact lens and apharmaceutical formulation of the invention.

In one aspect, the invention provides a kit containing a contact lensand a pharmaceutical formulation of the invention, in a container thatis unopened subsequent to manufacture.

In one aspect, the invention provides a kit containing a sterilepharmaceutical formulation of the invention and packaging materialstherefore which permit use as an artificial tear product introducibleinto the eye.

In one aspect, the invention provides a kit comprising an extraoculardevice coated with the composition of the invention, and packagingmaterials therefore. In various embodiments, the packaging materialspermit contacting of the extraocular device through the nozzle of an eyecare solution container. In various embodiments, the extraocular deviceis an intracorneal inlay.

In one aspect, the invention provides a method for manufacture of acontact lens, involving providing a contact lens and contacting thecontact lens with a composition of the invention.

In another aspect, the invention provides a kit containing a contactlens and a pharmaceutical formulation of the invention, in a containerthat is reusable.

In one aspect, the invention provides a copolymer coated nanoparticle,the surface of the nanoparticle containing a copolymer having apositively charged, or hydrophobic, or covalent bonding moiety and ahydrophilic moiety. In various embodiments, the nanoparticle furthercontains an active agent.

An aspect of the present invention relates to a method for changingwettability of and/or sterically modifying a biological surface in asubject comprising locally administering to the biological surface ofthe subject a pharmaceutical formulation comprising a bi-functionalco-polymer. A copolymer composition can be contacted with an externalbody surface (e.g., skin) and/or used to prevent infection. Ofparticular interest and focus in this invention is the external surfaceof the eye including bulbar and palpebral conjunctiva and the cornea.Another aspect of the present invention relates to a method for treatingdry eye syndrome comprising topically administering to an eye of asubject with dry eye syndrome a pharmaceutical formulation comprising abi-functional co-polymer.

Another aspect of the present invention relates to a method forprotecting a biological surface in a subject against pathogen or toxinattack, said method comprising topically or locally administering to thebiological surface in the subject a pharmaceutical formulationcomprising a bi-functional co-polymer.

Another aspect of the present invention relates to a method fordelivering an active pharmaceutical ingredient to a biological surfacein a subject comprising topically or locally administering to thebiological surface of the subject a pharmaceutical formulationcomprising a bi-functional co-polymer and the active pharmaceuticalingredient.

Another aspect of the present invention relates to a formulation for usein methods of changing wettability of (or otherwise sterically modifyingor stabilizing) a biological surface in a subject, treating orpreventing dry eye syndrome, treating or preventing discomfort relatedcomplications of LASIK surgery, treating or preventing infectious orinflammatory conditions of the external eye, protecting a biologicalsurface in a subject against pathogens (bacteria, fungi, virus, prions,bioadhesive chemical agents) and/or delivering an active pharmaceuticalingredient to a biological surface in a subject, said formulationcomprising a bi-functional co-polymer for topical and/or localadministration to the subject.

Another aspect of the present invention relates to coating extraoculardevices with a bi-functional co-polymer. Another aspect of the presentinvention relates to the use of the bi-functional co-polymers describedherein to prevent or treat contact lens related discomfort and to reducesymptoms of contact lens intolerance.

Other aspects of the invention include the coating and thus protectionof biological surfaces, such as the skin, especially mucous membranesinside the mouth, nose, throat, ear, vagina, for the purpose ofmaintaining hydration, and for protecting the biological surface frominfection by pathogens and from toxins.

Other aspects of the invention include coating drug particles orparticles containing drug to increase their utility by increasing halflife and/or decreasing interaction with biologic phenomenon such as animmune response.

In various embodiments of any of the above aspects, the copolymer is agraft or a block copolymer. In various embodiments, the copolymer is agraft copolymer and contains a cationic backbone and side chains thatare water soluble and non-ionic. In various embodiments, the copolymeris a graft copolymer and contains a water soluble non-ionic backbone andcationic side chains. In various embodiments, the copolymer is a blockcopolymer and contains at least one cationic block and at least onewater soluble and non-ionic block. In various embodiments, the copolymeris a block copolymer and contains at least one block which ishydrophobic and at least one block which is water soluble and anionic.In various embodiments, the copolymer is a block copolymer and containsat least one block which is hydrophobic and at least one block which iswater soluble and cationic. In various embodiments, the graft copolymeris PLL-g-PEG.

In various embodiments of any of the above aspects, the copolymer is adendrimer. In various embodiments of any of the above aspects, thehydrophilic moiety is one of non-ionic or anionic. In variousembodiments of any of the above aspects, the copolymer is a blockcopolymer and contains at least one block which is hydrophobic and atleast one block which is water soluble and or non-ionic.

In various embodiments of any of the above aspects, the copolymercreates a covalent adhesion between the copolymer and the biologicalsurface. In various embodiments of any of the above aspects, thecontacted biological surface is the surface of an eye, a mucousmembrane, or skin of a subject.

In various embodiments of any of the above aspects, the copolymer is0.001 to 40% of the composition. In various embodiments of any of theabove aspects, the copolymer is 0.001 to 25% of the composition.

In various embodiments of any of the above aspects, the compositionfurther contains a second polymer. In various embodiments of any of theabove aspects, the composition further contains a PLURONIC blockcopolymer. In various embodiments of any of the above aspects, thecomposition further contains one or more of a surfactant, apreservative, and a pharmaceutical ingredient including demulcents[Cellulose derivatives (Carboxymethylcellulose sodium, Hydroxyethylcellulose, Hydroxypropyl methylcellulose, Methylcellulose), Dextran 70,Gelatin, Polyols (Glycerin, Polyethylene glycol 300, Polyethylene glycol400, Polysorbate 80, Propylene glycol), Polyvinyl alcohol, Povidone]emollients (Lanolin, mineral oil, paraffin, petrolatum, white ointment,white petrolatum, white wax, yellow wax), sodium chloride, andvasoconstrictors (Ephedrine hydrochloride, Naphazoline hydrochloride,Phenylephrine hydrochloride, Tetrahydrozoline hydrochloride).

In various embodiments of any of the above aspects, the composition iscontacted to the biological surface in an amount sufficient to changetribological properties of the biological surface of the subject. Invarious embodiments of any of the above aspects, the composition iscontacted topically to an eye of a subject. In various embodiments ofany of the above aspects, the composition is topically administered toan eye of a subject to treat dry eye, ophthalmic irritation, or cornealepithelial disease. In various embodiments, the composition is contactedto the eye by an eye drop or an eye care solution. In variousembodiments, the method further involves administering to the eye of thesubject a second, different eye drop. In various embodiments of any ofthe above aspects, the composition has a low viscosity artificial tearless than 20 cP, having prolonged tear film break up time. In variousembodiments of any of the above aspects, the positively charged moietyis sufficient as a preservative in a topical ophthalmic formulation. Invarious embodiments of any of the above aspects, composition is in avolume sufficient for instillation in the eye, wherein the copolymer isPLL-g-PEG at a concentration of 0.1 to 3 wt %.

In various embodiments of any of the above aspects, the copolymer isimmobilized on an extraocular device at the time of manufacture. Invarious embodiments, the copolymer is immobilized on the extraoculardevice covalently. In various embodiments, the immobilization iseffected via an aldehyde plasma polymer interlayer and reductiveamination. In various embodiments, the extraocular device is a contactlens.

BRIEF DESCRIPTION OF THE FIGURES

FIG. 1 shows results from an in vitro assessment of the ability ofexemplary formulations of the present invention containing PLL-g-PEG todecrease contact angle of water to polystyrene. Formulations containing10 mg/ml or 1% (closed circles), 5 mg/ml or 0.5% (open circles) or 1mg/ml or 0.1% (closed triangles) were compared to a control of deionizedwater (open triangles).

FIG. 2 shows results from an in vitro assessment of the ability ofexemplary formulations of the present invention containing F87 or F127to decrease contact angle of water to polystyrene. Formulationscontained either 1% F127 (open circles) or 1% F87 (closed circles) andwere compared to the average contact angle of deionized water (straightline).

FIG. 3 shows the results of a randomized, controlled, double maskedclinical trial in sixteen subjects where the tear film break-up time wasmeasured at varying time points after eye drop instillation. Thisclinical methodology is a common assessment tool for dry eye syndrome,as well as to determine the effectiveness of artificial tears. Fifteenminutes after instillation of sample formulation and active control,sample formulation showed extension of noninvasive tear film break-uptime from baseline of 14.67 sec (p=0.05), while active comparator was7.4 seconds longer than baseline (p=0.34).

FIG. 4 shows results from the same randomized, controlled, double maskedclinical trial where the tear film break-up time was measured at varyingtime points in subjects. Fluorescein break-up time was longer thanactive comparator at 120 minutes; superior by 4.92 sec (p=0.12).

FIG. 5 shows results of a sub group analysis of the trial describedabove. In the main group of 16 subjects in the trial, there were threesubjects that had unexpectedly long pre-instillation TFBUT. These longinitial values distorted the relative contributions of the artificialtears to NIBUT for the other subjects. When these three outliers wereremoved, there was superiority in extending NIBUT for the sampleformulation at every time point. The data is graphed based on extensionby seconds over baseline in FIG. 5.

FIG. 6 shows results of a sub group analysis of the trial describedabove similar in method to FIG. 5. In the main group of 16 subjects inthe trial, there were three subjects that had unexpectedly longpre-instillation TFBUT. These long initial values distorted the relativecontributions of the artificial tears to NIBUT for the other subjects.When these three outliers were removed, there was superiority inextending NIBUT for the sample formulation at every time point. The datais graphed by percentage change from baseline in FIG. 6.

FIG. 7 is an example schematic drawing of the molecular behavior of thegraft copolymer on an epithelial or ophthalmic device surface. Thisschematic demonstrates how the surface modifying polymer can bebeneficial in altering the wettability or tribological properties of asurface and/or sterically modifying the surface so that the epitheliumis protected and stays wet longer, or how an extra- or intra-oculardevice may be made more biocompatible and less likely to foul withproteins and cellular debris. 701 represents the negatively chargedepithelial surface (such as the cornea and conjunctiva) or negativecharges on a device (such as a contact lens). 702 represents thepositive charge on the NH3 terminal of lysine. 703 represents theanchoring poly (L) lysine backbone. 704 represents the flexiblepolyethylene glycol (PEG) side chains that form a brush. This brush canimpart protein resistance, and is hydrated (705 is a water molecule),and thus there is also a change in wettability imparted onto the surfaceon which the graft (or block) copolymer electrostatically (or throughhydrophobic interaction in some cases) adheres. 706 represents theentire graft copolymer molecule (PLL-g-PEG in this example). Stericstabilization is also imparted to exposed surfaces decreasing contactwith proteins and cells, thereby protecting the epithelial or devicesurface from protein or cellular adhesion, inflammation or fouling. Thisfigure is an example of PLL-g-PEG, but the same concept applies forother graft copolymers, generally, using either hydrophobic or cationicinteraction with a surface combined with a hydrophilic moiety impartingthe protective aspects of the invention herein described. Furthermore,the bi-functional co-polymers, block copolymers, and dendrimers may bediagrammed in a similar fashion, using a repeating moiety adherent tothe surface and a repeating hydrophilic moiety imparting changes inwettability and protection. Furthermore, in some embodiments, thebi-functional copolymer may be covalently immobilized on the surface ofa device such as a contact lens.

DETAILED DESCRIPTION OF THE INVENTION

It has now been found that topical or local administration of aformulation comprising a graft co-polymer having a positively charged orhydrophobic moiety and a hydrophilic moiety or a block co-polymer havinga positively charged or hydrophobic moiety and a hydrophilic moiety to abiological surface alters and increases the wettability of certainsurfaces, including a biological surface particularly the corneaconjunctiva and external surface of the eye, and does so for a prolongedperiod of time. A copolymer composition of the invention can becontacted with an external body surface (e.g., skin) and/or used toprevent infection.

By “moiety” it is meant the required portion of the polymer. Forexample, the moiety may be a monomer or one aspect of the polymer thatimparts some characteristic required for activity and/or result. The PEGmoiety imparts hydrophilicity. The PLL moiety imparts cationic charges.A phenylboronic moiety for example imparts an ability to form covalentattractions.

By “covalent” it is meant an interatomic bond characterised by thesharing of electrons. The bond can be reversible or permanent.

By “electrostatic” it is meant interaction between two molecules or twopolymers or between a molecule or polymer and a biological ornon-biological surface based on charge attractions—negative charge onone aspect of entity and positive charge on one aspect of the other.

By “bioadhesive” is meant a synthetic or natural polymer which binds tobiological substrates such as mucosal membranes. Binding may beaccomplished by electrostatic interaction, covalent attraction,hydrophobic interaction or other means.

By “hydrophobic interaction” it is meant an interaction betweenlipophilic moieties on polymers and surfaces to form adhesions orintermolecular aggregates or intramolecular interactions (particularlyin aqueous based environments).

By “adhesive”, it is meant tending to adhere, stick, or not come easilyloose, or tending to persist.

By “increases the wettability” it is meant an increase in the relativedegree with which an aqueous liquid will spread onto or coat a surface,including in the presence of other immiscible liquids. Increasedwettability, according to the invention results in prolonged stabilityof the tear film keeping the cornea better coated by natural andartificial tear film components, reduce adhesion of inflammatoryproteins, bacteria and inflammatory proteins to a biological surface ormembrane of the subject.

Changes in wettability, however, are only part of the advantagesconveyed by this invention. Surface modifications also include thesteric stabilization of biological surfaces and extra-ocular ophthalmicdevice surfaces.

By “biological surface” is meant the surface of a bodily organ, whetherit be exposed to the external environment, or internal to the body. Forexample, the surface of the eye includes the epithelial covered corneaand bulbar and palpebral conjunctiva, as well as the posterior tenonslayer and sclera; the epithelial layers of the gastrointestinal tract orthe skin are included, membranes such as mucous membranes, includingoral, nasal, respiratory, urinary tract including the bladder, andvaginal mucous membranes. Other surfaces include the capsules of organssuch as the spleen and liver, and the outermost aspect of bone,cartilage, and muscle, as well as wounds and areas where there isunprotected subepithelial biological tissue exposure.

By “tribological properties” it is meant properties of interaction ofsurfaces in relative motion. For example, a contact lens on the surfaceof the eye with some movement on the corneal surface and/or with the actof blinking whereby the palpebral conjunctival surface moves over thecontact lens constitutes a state of relative motion between twosurfaces. Likewise, blinking whereby the palpebral conjunctival surfacemoves over the cornea and bulbar conjunctiva constitutes a state ofrelative motion between two surfaces. Tribological properties caninclude but are not limited to properties such as lubrication, friction,and wear.

By “subject”, as used herein it is meant to be inclusive of all animalsand in particular mammals such as, but not limited to, humans and dogsas well as agricultural animals such as bovine, ovine, and porcine.

By “prolonged period of time” means that the formulation has an effectthat lasts longer than leading artificial tear products on the market,which has been shown to be at least two hours with the sampleformulation, but “prolonged period of time” can also mean longer than 20minutes, as most ophthalmic topical products are washed out of the eyein 20 minutes. A “prolonged” period of time includes 30, 45, 50 minutes,as well as 1, 2, 5, 10, 24 hours, or 1, 2, 5, 10 or 30 days or more.

By “steric stabilization” or “steric modification” means that thesurface effects of the graft and block co-polymers results in ahydrophilic moiety projected away from the biological surface, drugparticle or drug containing particle that imparts characteristics suchas low protein adsorption and cell adhesion. Such an effect can also becalled steric repulsion and surface exclusion effects secondary to thePEG moieties (termed “brushes” in some descriptions) on the graftcopolymers. Other hydrophilic chains can accomplish the same purpose insome embodiments. High surface water retaining capacities, osmoticrepulsion, and charge neutrality may also contribute to this “stericstabilization” effect.

By “storage” is meant time on the shelf in a container, and storage timeincludes 1, 2, 5, 10, 30, 180, or 360 days or more.

Accordingly, the present invention provides methods for changingwettability of (or sterically stabilizing or otherwise modifying)biological surfaces or membranes in a subject via topical or localadministration of a pharmaceutical formulation comprising a graftco-polymer having a positively charged or hydrophobic moiety and ahydrophilic moiety or a block co-polymer having a positively charged orhydrophobic moiety and a hydrophilic moiety to a biological surface of asubject, or both. Pharmaceutical formulations can be administeredtopically or locally in accordance with methods of the present inventionto biological surfaces of a subject including, but not limited to, skin,mucous membranes, hair, and the surface of the eye.

According to the present invention, through the use of the block andgraft co-polymers described herein, certain drugs and activebiopharmaceutical agents will have improved efficacy and decreasedfrequency of dosing needs as formulations using the block or graftcopolymers can bring such active pharmaceutical agents into contact withepithelial surfaces, such as the cornea and conjunctiva, for prolongedperiods of time.

According to the present invention, through the use of the block andgraft co-polymers described herein, the tolerability of extraoculardevices such as contact lenses, shunts, retinal implant/explantmaterials and devices for scleral buckles, and drug delivery devices,can be improved. Furthermore, ophthalmic sutures may be coated withthese polymers decreasing tissue reaction and reducing unwanted particleor bacterial adherence. First, the devices can be coated (usingelectrostatic or hydrophobic interactions or through covalentimmobilization), with the said polymers increasing wettability,modifying surface characteristics, and/or sterically stabilizing anddecreasing the adherence of unwanted antigens, cells, proteins, andother particles to them. Second, by protecting the epithelial surface orother ophthalmic tissues (through turnover and/or exchange of theseadherent polymers with a biological surface and/or through theseparation of the device from the epithelial surface by the “brush”characteristics of the hydrophilic moieties), the devices may be moretolerable.

By “extraocular device” is meant medical devices that do touch but donot reside entirely inside the eye. A contact lens is an extraoculardevice. An intra-corneal inlay is an extraocular device because it doesnot breach the intraocular environment. A glaucoma shunt, is anextraocular device because the vast majority of the device (except forthe distal tip of the tube) resides outside the eye. A scleral buckle isan extraocular device.

By “co-polymers” is meant a polymer with two or more different monomers.Block, graft, and dendrimer co-polymers are the major, but not onlytypes of co-polymers referenced herein.

By “bi-functional” is meant the co-polymer has two or more differentfunctions that derive from the different moieties or monomers inherentto the co-polymer. For example a hydrophilic moiety or monomer has theproperty of wettability and a cationic backbone has the property ofbioadhesiviness. Joined together in a co-polymer both functions remain.Specifically, bifunctional co-polymers relates to but is not limited tograft, block, a and dendrimer co-polymers. Critical to this invention isthe novel use of bi-functional graft copolymers in ophthalmic use.

By “ophthalmic use” is meant used on or in or adjacent to the eye.Topical eye drops are included in ophthalmic use as is subconjunctival,peribulbar and eyelid use. Use on contact lenses and extraocular devicesis included in ophthalmic use.

By “pluronic” is meant poloxamers or trade name pluronic copolymers thatare composed of nonionic triblock copolymers composed of a centralhydrophobic chain of polyoxypropylene (poly(propylene oxide)) flanked bytwo hydrophilic chains of polyoxyethylene (poly(ethylene oxide))

By “preserved” it is meant there is a substance or preparation added toa product to destroy or inhibit the multiplication of microorganisms.

According to the present invention, through the use of the block andgraft co-polymers described herein, the tolerability of extraoculardevices (such as contact lenses and intracorneal optical devices), canbe improved. First, the devices can be coated with the said polymersincreasing wettability and decreasing the adherence of unwantedantigens, cells, proteins, and other particles to them. Second, byprotecting the adjacent tissue structures, the devices may be moretolerable and function better i.e. through less protein or cellularinteraction on the device. In the case of intracorneal inlay technology,such a coating may improve the clinical performance of such devices inmultiple ways.

Graft co-polymers used in the methods and formulations of the presentinvention are polymers having a linear section of repeat units calledthe “backbone”, with at least one side chain of repeat units (called agraft), usually of a different chemistry, branching from a point alongthe backbone. In one embodiment, the graft co-polymer comprises acationic backbone and side chains that are water soluble and non-ionic.In another embodiment, the graft copolymer comprises a water solublenon-ionic backbone and cationic side chains.

Block co-polymers used in the methods and formulations of the presentinvention are polymers in which linear sections of a first section ofrepeat units are connected end-to-end with linear sections of subsequentrepeat units that are chemically dissimilar to the first.

Dendrimers comprised of cationic and hydrophilic polymeric moieties cansimilarly be used as identified above in the methods and formulations ofthe present invention to accomplish similar structural and stericalterations of the biological surface (particularly the eye), drugparticles or drug containing particles.

By “dendrimers” it is meant co-polymer molecules that are repeatedlybranched.

Also included in the invention is the use of graft, block, anddendrimeric copolymers with multi-functionality in topical applicationsand formulations including ophthalmic applications and formulations. Forexample, a polymer where there is a moiety bonded to one constituentpolymer that promotes covalent bond formation between the copolymer andbiological surfaces such as the surface of the eye or between thecopolymer and extraocular devices that also project a comb or a brushtype of hydrophilic moiety, is claimed. As can be seen to one skilled inthe art, the results accomplished with the use of electrostaticinteraction between the polymer and biological surfaces and the externalenvironment or extraocular devices can be accomplished using similarlydesigned bi- and multi-functional polymers that promote the formation ofa covalent bond instead of or in addition to electrostatic attractiononly. For example, the use of phenylboronic acid (PBA) or other boronbased moieties in the polymer backbone provide a method for theformation of covalent complexes between copolymers and biologicalsurfaces and between copolymers and extraocular devices. Even though notcalled out continuously throughout this document, the method of usingcovalent, electrostatic and hydrophobic attraction (or any combinationor either) in ophthalmic applications is described. PLL-g=(PEG;PBA) isan example of a polymeric structure that would impart the describedcharacteristics.

Formulations for use in the methods of the present invention comprise ablock or graft co-polymer having one section, either the backbone, thegraft or the block, that adheres to a biological surface tissue such as,but not limited to, the eye surface by electrostatic or hydrophobicforces (or covalent), and another, chemically different section, eitherthe backbone, the graft, or block, that is hydrophilic and retainsmoisture (sometimes called a “brush” or a “PEG brush” in someembodiments), or allows an aqueous film to readily spread over and thuswet the biological surface. Sometimes the graft copolymers such asPLL-g-PEG, for example, can be referred to as comb co-polymers.Furthermore, the “brush” that is composed of water soluble polymers thatare biocompatible can in aqueous environment, provide oil free lubricity(Drobeck et al. Langmuir. 24(4):1484-8).

The surface modifications imparted by graft and block co-polymers hereindescribed may include, but are not limited to changes in wettability,steric modification or steric stabilization, steric repulsion, surfaceexclusion effects, high surface water retaining characteristics, chargeneutrality, and osmotic repulsion. These and other effects on thesurfaces are important processes that may convey clinically meaningfulbenefits to subjects and to the performance of medical devices. Thetissue-adhesive sections of a bi-functional co-polymer in theformulations used in the methods of the present invention may becationic, in which case the polymer adheres to the biological surface byelectrostatic attraction, or may be hydrophobic in which case thepolymer adheres to the biological surface by hydrophobic interaction, ormay involve a covalent complex. In some cases, the wettability of thebiological surface may result in whole or in part from modification of aliquid layer adjacent the surface, such as the mucous layer adjacent thecornea. In some cases the tissue surface modification may simply imparta lubricious protective coating over ophthalmic tissue or extraoculardevices. It may also alter the tribological properties of tissues andextraocular devices.

Aspects of the invention include copolymer coated nanoparticles whichcan encompass active agents, including drugs (small molecule, chemical,pharmaceutical, biologic) or allergens (viruses, bacteria, yeast,prions). The nanoparticles have exposed on their surface (i.e., arecoated with) a copolymer, where the copolymer has a positively charged,or hydrophobic, or covalent bonding moiety and a hydrophilic moiety.Nanoparticles are not of sufficient size to encompass cells.Nanoparticles are not microspheres. Other aspects of the inventioninclude coating drug particles or particles containing drug to increasetheir utility by increasing half life and/or decreasing interaction withbiologic phenomenon such as an immune response. For intramuscularinjection, the nanoparticle size should be 1 mm or smaller diameter; forinhalation, the nanoparticle size should be 10 microns or smaller indiameter; for IV injection, the nanoparticle size should be 1 micron orsmaller in diameter.

Aspects of the invention include copolymer coated nanoparticles whichcan encompass active agents, including drugs (small molecule, chemical,pharmaceutical, biologic) or allergens (viruses, bacteria, yeast,prions). The nanoparticles have exposed on their surface (i.e., arecoated with) a copolymer, where the copolymer has a positively charged,or hydrophobic, or covalent bonding moiety and a hydrophilic moiety.Nanoparticles are not of sufficient size to encompass cells.Nanoparticles are not microspheres. Other aspects of the inventioninclude coating drug particles or particles containing drug to increasetheir utility by increasing half life and/or decreasing interaction withbiologic phenomenon such as an immune response.

Nanoparticles of the invention have an effective average particle sizeof less than about 1000 nm (i.e., 1 micron), less than about 900 nm,less than about 800 nm, less than about 700 nm, less than about 600 nm,less than about 500 nm, less than about 400 nm, less than about 300 nm,less than about 250 nm, less than about 200 nm, less than about 150 nm,less than about 100 nm, less than about 75 nm, or less than about 50 nm,as measured by light-scattering methods, microscopy, or otherappropriate methods. By “an effective average particle size of less thanabout 2000 nm” it is meant that at least 50% by weight of the activeagent particles have a particle size less than the effective average,i.e., less than about 2000 nm, 1900 nm, 1800 nm, etc., when measured bythe above-noted techniques. In other embodiments of the invention, atleast about 70%, at least about 90%, or at least about 95% of the activeagent particles have a particle size less than the effective average,i.e., less than about 2000 nm, 1900 nm, 1800 nm, etc. For intramuscularinjection, the nanoparticle size should be 1 mm or smaller diameter; forinhalation, the nanoparticle size should be 10 microns or smaller indiameter; for IV injection, the nanoparticle size should be 1 micron orsmaller in diameter.

Nanoparticles of the invention can be made using, for example, milling,homogenization, or precipitation techniques. Exemplary methods of makingnanoparticulate compositions are described in the '684 patent. Methodsof making nanoparticulate compositions are also described in U.S. Pat.No. 5,518,187 for “Method of Grinding Pharmaceutical Substances;” U.S.Pat. No. 5,718,388 for “Continuous Method of Grinding PharmaceuticalSubstances;” U.S. Pat. No. 5,862,999 for “Method of GrindingPharmaceutical Substances;” U.S. Pat. No. 5,665,331 for“Co-Microprecipitation of Nanoparticulate Pharmaceutical Agents withCrystal Growth Modifiers;” U.S. Pat. No. 5,662,883 for“Co-Microprecipitation of Nanoparticulate Pharmaceutical Agents withCrystal Growth Modifiers;” U.S. Pat. No. 5,560,932 for“Microprecipitation of Nanoparticulate Pharmaceutical Agents;” U.S. Pat.No. 5,543,133 for “Process of Preparing X-Ray Contrast CompositionsContaining Nanoparticles;” U.S. Pat. No. 5,534,270 for “Method ofPreparing Stable Drug Nanoparticles;” U.S. Pat. No. 5,510,118 for“Process of Preparing Therapeutic Compositions ContainingNanoparticles;” and U.S. Pat. No. 5,470,583 for “Method of PreparingNanoparticle Compositions Containing Charged Phospholipids to ReduceAggregation,” all of which are herein incorporated by reference.

The resultant nanoparticulate active agent compositions can be utilizedin solid or liquid dosage formulations, such as controlled releaseformulations, solid dose fast melt formulations, aerosol formulations,nasal formulations, lyophilized formulations, tablets, capsules, solidlozenge, powders, creams, ointments, etc.

The nanoparticles of the invention can be contacted with a copolymerhaving a positively charged, or hydrophobic or covalent bonding moietyand a hydrophilic moiety after attrition. One or more secondary surfacestabilizers may also be added before or after attrition. The activeagent particles can be reduced in size in the presence of a copolymerhaving a positively charged, or hydrophobic or covalent bonding moietyand a hydrophilic moiety. Other compounds, such as a diluent, can beadded to the copolymer active agent/surface stabilizer compositionduring the size reduction process (e.g., milling). Dispersions can bemanufactured continuously or in a batch mode.

Another method of forming the copolymer coated nanoparticles of theinvention is by microprecipitation. This is a method of preparing stabledispersions of copolymers or active agents in the presence of one ormore surface stabilizers and one or more colloid stability enhancingsurface active agents free of any trace toxic solvents or solubilizedheavy metal impurities. Such a method comprises, for example: (1)dissolving the copolymer or active agent in a suitable solvent; (2)adding the formulation from step (1) to a solution containing acopolymer having a positively charged, or hydrophobic or covalentbonding moiety and a hydrophilic moiety and/or active agent andoptionally one or more secondary surface stabilizers, to form a clearsolution; and (3) precipitating the formulation from step (2) using anappropriate non-solvent. The method can be followed by removal of anyformed salt, if present, by dialysis or diafiltration and concentrationof the dispersion by conventional means.

Exemplary homogenization methods of preparing copolymer coatednanoparticles are described in U.S. Pat. No. 5,510,118, for “Process ofPreparing Therapeutic Compositions Containing Nanoparticles.” Such amethod comprises dispersing nanoparticles in a liquid dispersion mediumin which the copolymer or active agent is soluble, followed bysubjecting the dispersion to homogenization to reduce the particle sizeof the active agent to the desired effective average particle size. Thenanoparticles can be reduced in size in the presence of a copolymerhaving a positively charged, or hydrophobic or covalent bonding moietyand a hydrophilic moiety and/or active agent, and, if desired, one ormore additional surface stabilizers. Alternatively, the nanoparticlescan be contacted with the copolymer and/or active agent and, if desired,one or more additional surface stabilizers either before or afterattrition. Other compounds, such as a diluent, can be added to theactive agent/copolymer composition either before, during, or after thesize reduction process. Dispersions can be manufactured continuously orin a batch mode.

The copolymer coated nanoparticles of the invention may contain anactive agent. The active agent may be contained within the nanoparticle,on the surface of the nanoparticle, in the copolymer coating of thenanoparticle, or on the surface of the copolymer coating of thenanoparticle. Active agents that can be used with the copolymer coatednanoparticle include without limitation drugs (small molecule, chemical,pharmaceutical, biologic) or allergens (viruses, bacteria, yeast,prions). Active agents may be therapeutic, or diagnostic agent. Atherapeutic agent can be a pharmaceutical agent, including biologicssuch as proteins, peptides, and nucleotides, or a diagnostic agent, suchas a contrast agent, including x-ray contrast agents. The active agentexists either as a discrete, crystalline phase, an amorphous phase, asemi-amorphous phase, a semi-crystalline phase, or mixtures thereof. Thecrystalline phase differs from a non-crystalline or amorphous phasewhich results from precipitation techniques, such as those described inEP Pat. No. 275,796. Two or more active agents can be used incombination.

The active agent can be selected from a variety of known classes ofdrugs, including, for example, proteins, peptides, nucleotides,anti-obesity drugs, nutraceuticals, dietary supplements, carotenoids,corticosteroids, elastase inhibitors, anti-fungals, oncology therapies,anti-emetics, analgesics, cardiovascular agents, anti-inflammatoryagents, anthelmintics, anti-arrhythmic agents, antibiotics (includingpenicillins), anticoagulants, antidepressants, antidiabetic agents,antiepileptics, antihistamines, antihypertensive agents, antimuscarinicagents, antimycobacterial agents, antineoplastic agents,immunosuppressants, antithyroid agents, antiviral agents, anxiolyticsedatives (hypnotics and neuroleptics), astringents, beta-adrenoceptorblocking agents, blood products and substitutes, cardiac inotropicagents, contrast media, corticosteroids, cough suppressants(expectorants and mucolytics), diagnostic agents, diagnostic imagingagents, diuretics, dopaminergics (antiparkinsonian agents),haemostatics, immunological agents, lipid regulating agents, musclerelaxants, parasympathomimetics, parathyroid calcitonin andbiphosphonates, prostaglandins, radio-pharmaceuticals, sex hormones(including steroids), anti-allergic agents, stimulants and anoretics,sympathomimetics, thyroid agents, vasodilators, and xanthines.

Exemplary nutraceuticals and dietary supplements are disclosed, forexample, in Roberts et al., Nutraceuticals: The Complete Encyclopedia ofSupplements, Herbs, Vitamins, and Healing Foods (American NutraceuticalAssociation, 2001), which is specifically incorporated by reference. Anutraceutical or dietary supplement, also known as phytochemicals orfunctional foods, is generally any one of a class of dietarysupplements, vitamins, minerals, herbs, or healing foods that havemedical or pharmaceutical effects on the body. Exemplary nutraceuticalsor dietary supplements include, but are not limited to, lutein, folicacid, fatty acids (e.g., DHA and ARA), fruit and vegetable extracts,vitamin and, mineral supplements, phosphatidylserine, lipoic acid,melatonin, glucosamine/chondroitin, Aloe Vera, Guggul, glutamine, aminoacids (e.g., iso-leucine, leucine, lysine, methionine, phenylanine,threonine, tryptophan, and valine), green tea, lycopene, whole foods,food additives, herbs, phytonutrients, antioxidants, flavonoidconstituents of fruits, evening primrose oil, flax seeds, fish andmarine animal oils, and probiotics. Nutraceuticals and dietarysupplements also include bio-engineered foods genetically engineered tohave a desired property, also known as “pharmafoods.”

Active agents to be administered in an aerosol formulation arepreferably selected from the group consisting of proteins, peptide,bronchodilators, corticosteroids, elastase inhibitors, analgesics,anti-fungals, cystic-fibrosis therapies, asthma therapies, emphysematherapies, respiratory distress syndrome therapies, chronic bronchitistherapies, chronic obstructive pulmonary disease therapies,organ-transplant rejection therapies, therapies for tuberculosis andother infections of the lung, fungal infection therapies, respiratoryillness therapies associated with acquired immune deficiency syndrome,an oncology drug, an anti-emetic, an analgesic, and a cardiovascularagent.

A description of these classes of active agents and a listing of specieswithin each class can be found in Martindale, The Extra Pharmacopoeia,Twenty-ninth Edition (The Pharmaceutical Press, London, 1989),specifically incorporated by reference. The active agents arecommercially available and/or can be prepared by techniques known in theart.

Exemplary active agents to be applied to mucous include dentalapplications, such as oral bioadhesive nanoparticulate lidocainformulations, bioadhesive nanoparticulate fluoride treatments,application to the lungs, throat, GIT, application to wounds, etc. Alsoincluded is application to the throat using a liquid containing abioadhesive nanoparticulate formulation containing, for example, mentholor other numbing compound for treatment of coughs or sore throats. Thestomach and GIT can also be treated using bioadhesive formulations. Thisis particularly useful for treatment of diseases associated with themucous of the gastrointestinal tract, such as Crohn's Disease. Otherpharmaceutical therapeutic methodologies include oral dosing, nasaladministration, vaginal administration, ocular administration, colonic,and subcutaneous administration.

The compositions of the invention also encompass food products. Forexample, spice, oleoresin, flavor oil, color, or chemicals are oftenadded during food processing to produce the desirable flavors, taste,and appearance. These agents can be included in a bioadhesivenanoparticulate composition of the present invention for increasedadhesion to biological surfaces. Bioadhesive nanoparticulate flavoringagents could be used in products such as gums to produce prolongedflavor.

Other exemplary uses of the novel bioadhesive formulations are provided:teeth can be treated with teeth whiteners or fluoride bioadhesivecompositions; bones can be treated with calcium bioadhesivecompositions; nails can be treated with color or strengtheningbioadhesive formulations; insects or pests can be treated withinsecticides or other toxic compositions to the pest. In sum, thecompositions are useful in treating any biological surface, or a surfacederived from a biological material. Feathers and scales of animals canbe treated, as well as other animal biological surfaces such as chitin.

For the purpose of this invention, the description that follows below onthe different types of graft and block co-polymers applies to inventionembodiments of formulations for ophthalmic and medical topical andpharmaceutical use, and to the use of these graft and block co-polymersin the setting of extra- and intra-corneal ophthalmic devices ascoatings, and for ways of treating, storing, or manufacturing, orreapplying the coatings to these devices:

Examples of cationic polymer sections of graft or block co-polymers offormulations useful in the methods of the present invention include, butare not limited to: polyp lysine) (PLL), polylysine [includingpoly-D-lysine (PDL), poly-DL-lysine, poly-e-CBZ-D-lysine,poly-e-CBZ-DL-lysine, or poly-e-CBZ-L-lysine) polyornithine (i.e.,poly-DL-ornithine, poly-L-ornithine or poly-S-CBZ-DL-ornithine), and thelike)], [note Pub. No.: WO/1993/018649 International Application No.:PCT/US1993/002609 contains additional descriptions of polycatioinc watersoluble graft copolymers and is incorporated here by reference],poly(2-vinyl pyridine, poly(4-vinyl pyridine) and vinyl co-polymerscontaining those repeat units, and poly(aminoethyl methacrylate) homo-and co-polymers containing N,N dimethylaminoethylmethacrylate) repeatunits. In general, polymers containing acrylates and acryalmides canserve as cationic sections of the bi-functional co-polymers. Additionalexemplary cationic polymers include, but are not limited to,poly(trimethylammonioethyl methacrylate chloride),poly-(2-(dimethylamino) ethyl methacrylate) (pDMAEMA),poly-(2-(dimethylamino) ethyl methacrylateco-methacrylic acid)(pDMAEMA-co-MAA) and poly-(2-methyl-acrylic acid2-[(2-dimethylamino-ethyl)-methylamino]-ethyl ester) (pDAMA), and cationguar gum. Another cationic polymer section which can be used is chitosan(a co-polymer of glucosamine and N-acetyl glucosamine where 5-100% ofthe repeat units are glucosamine) and synthetic derivatives thereof. Theuse of amines and other positively charged amino acids bound topolymeric configurations to mimic the cationic polymeric activities sodescribed. In general, molecular structures that can impart a positiveexternal charge on a block or graft copolymer or a dendrimer areincluded in the invention.

Examples of hydrophobic polymer sections of bi-functional co-polymers offormulations useful in the methods of the present invention include, butare not limited to, long-chain aliphatic hydrocarbons, polyethylene,poly(propylene oxide), polystyrene, poly(methylmethacrylate),poly(butylenes oxide), and the like. In general, molecular structuresthat can impart an exposed hydrophobic domain on a block or graftcopolymer are included in the invention.

The hydrophilic section of the polymer may be non-ionic if the tissueadhering section is cationic, or non ionic or anionic if thetissue-adhering section is non-ionic (and hydrophobic). Examples ofnon-ionic hydrophilic polymer sections of formulations useful in methodsof the present invention include, but are not limited to,poly(ethyleneglycol) (PEG), poly(vinylalcohol),poly(vinylpyrrolidinone), dextrans and the like. Examples of anionichydrophilic polymer sections include homopolymers and co-polymerscontaining, for example, acrylic acid, methacrylic acid, itaconic acid,maleic acid, styrene sulfonic acid, carboxymethylcellulose,carboxyethylcellulose, succinylated chitosan, cellulose sulfate, and thelike. Additional exemplary hydrophilic polymers include, but are notlimited to, poly(dimethylamino)ethyl methacrylate and poly hydroxypropylmethacrylate (PHPMA).

The adsorbing segment could also comprise a boronate group selected fromthe group consisting of phenylboronic acid (PBA), 2-carboxyethaneboronicacid, 1,2-dicarboxyethaneboronic acid,.beta.,.beta.′-dicarboxyethaneboronate,.beta.,.gamma.-dicarboxypropaneboronate, 2-nitro- and4-nitro-3-succinamidobenzene boronic acids,3-nitro-4-(6-aminohexylamido)phenyl boronic acid,{4-[(hexamethylenetetramine)methyl]phenyl}boronic acid,4-(N-methyl)carboxamidobenzene boronic acid,2-{[(4-boronphenyl)methyl]-ethylammonio}ethyl and2-{[(4-boronphenyl)methyl]diethylammonio}-ethyl groups,succinyl-3-aminophenylboronic add, 6-aminocaproyl-3-aminophenylboronicacid, 3-(N-succinimidoxycarbonyl)aminophenylboronate,p-(.omega.-aminoethyl)phenylboronate, p-vinylbenzeneboronate,N-(3-dihydroxyborylphenyl)succinamic acid,N-(4-nitro-3-dihydroxyborylphenyl)succinamic acid,O-dimethylaminomethyl)benzeneboronic acid, 4-carboxybenzeneboronic acid,4-(N-octyl)carboxamidobenzeneboronic acid,3-nitro-4-carboxybenzeneboronic acid, 2-nitro-4-carboxybenzeneboronicacid, 4-bromophenylboronate, p-vinylbenzene boronate,4-(.omega.-aminoethyl)phenylboronate, catechol[2-(diethylamino)carbonyl,4-bromomethyl]phenyl boronate, and5-vinyl-2-dimethylaminomethylbenzeneboronic acid.

Adhesion resistant segments is another way to describe an importantcomponent of the copolymer. Adhesion resistant moieties could include:polyalkylene oxides, mixed polyalkylene oxides, polysaccharides,polyvinyl alcohol, poly-N-vinyl pyrrolidone, noncationic polyacrylates,noncationic polymethacrylates, and mixtures and copolymers of theseconstituents.

In general, the chains and block and graft copolymer moieties may be anylength, size, or molecular weight, and may have any number of repeatrates of the polymer components. Imparting beneficial properties is themain variable that will determine specific size, weights, chain lengths,and repeat specifications for applications described herein. As anexample, PEG of different molecular weights can be grafted onto thepoly-(L-lysine) at different ratios of PLL:PEG, to optimize the polymerarchitecture for wettability, steric stabilization, and proteinresistance. Additionally, other factors, such as cost, processdevelopment requirements, manufacturing, and biologic tolerability willgo into making specific size, weight, chain length, and repeatcharacteristic specification decisions. For these polymers,polydispersity may be acceptable over both wide and narrow ranges, andproduct specifications will be made based on this characteristic aswell. However, this invention claims the use of all sizes, weights,chain lengths, and repeat characteristics of these graft and blockco-polymers for uses and methods described herein.

Graft co-polymers may have a cationic (or non-ionic hydrophobic)backbone made from a polymer chosen from the list, supra, andhydrophilic grafts, or have a hydrophilic backbone, and cationic (ornon-ionic hydrophobic) grafts chosen from the list, supra. For graftco-polymers, grafts may arise from every repeat unit in the backbone ormay be intermittently spaced along the backbone (with uniform or randomfrequency). For example, a useful polymer in formulations for use in themethods of the present invention is PLL-g-PEG where the backbone is thecationic polymer poly(L-lysine) and the grafts are made from thehydrophilic polymer poly(ethylene glycol). The PLL backbone may be from3 repeat units to several thousand repeat units long, and the PEG graftsmay be from 1 to several thousand repeat units long. The PEG grafts maybe attached to every PLL repeat unit, every other PLL repeat unit, everythird repeat unit or less frequent. In one embodiment, there is a PEGgraft on average at every third PLL repeat unit. Regarding PLL, the PLLchain length can be any length, and made from PLL derived fromfermentation processes or synthetic polymerization reactions. Themolecular weight of the PLL backbone can be in the range of severalthousand Daltons (3,000 to 5,000) to tens of thousands of Daltons 15,000to 30,000, or higher. The chain length can be as short as ten lysinegroups or as long as one to two hundred (or longer) lysine groups. Thepolyamino acid chain bonds may be epsilon or alpha based. By block orgraft co-polymers it is meant to describe the architecture of thepolymer.

An additional exemplary polymer for use in formulations used in themethods of the present invention is PLL-g-dextran. Similar detailedanalysis and variability descriptions (as in the paragraph above) can bemade for PLL-g-dextran, and for many other graft co-polymers claimed inthis invention. Those skilled in the art will identify methods to mimicthe behavior of the copolymers herein described using different monomersand moieties, and those methods and compositions are claimed.

Block co-polymers or dendrimers comprising at least one block or moietythat is cationic and at least one block or moiety that is water solubleand non-ionic are also useful in formulations for use in methods of thepresent invention. In one embodiment, the co-polymer comprises at leastone block which is hydrophobic and at least one block which is watersoluble and anionic, cationic or non-ionic.

Examples of water soluble non-ionic co-polymer blocks or moietiesinclude, but are not limited to, poly(ethylene glycol) (PEG), polyvinylalcohol (PVA), poly(hydroxyethyl methacrylate) (pHEMA),poly(acrylamide), poly (vinyl pyrrolidone) (PVP), poly(ethyl oxazoline)(PEOX), polysaccharides, and copolymers of any two or more thereof.

Examples of water soluble anionic co-polymer blocks or moieties include,but are not limited to, polyacrylic acid (PAA), polymethacrylic acid,poly(sodium styrene sulfonate), carboxylated cellulosics such ascarboxymethylcellulose (CMC), poly(itaconic acid), poly(maleic acid),poly(acrylamidopropanesulfonic acid), anionic natural gums, anioniccarbohydrates, carageenan, alginates and hyaluronic acid.

Examples of water soluble cationic co-polymer blocks include, but arenot limited to, polymers based on vinyl pyridine,N,N-dimethylaminoethylacrylate, N,N-dimethylaminoethylmethacrylate,other acrylate and acryamide polymeric structures, allyl tri(alkyl)ammonium halides, poly(amino styrene), certain types of cation guargums, chitosan, polyethyleneimine, polyallylamine, polyetheramine,polyvinylpyridine, polysaccharides having a positively chargedfunctionality thereon, polyamino acids such as, but not limited to,poly-L-histidine, poly-im-benzyl-L-histidine, poly-D-lysine,poly-DL-lysine, poly-L-lysine, poly-ε-CBZ-D-lysine,poly-ε-CBZ-DL-lysine, poly-ε-CBZ-L-lysine, poly-DL-ornithine,poly-L-ornithine, poly-Δ-CBZ-DL-ornithine, poly-L-arginine,poly-DL-alanine-poly-L-lysine, poly(-L-histidine, L-glutamicacid)-poly-DL-alanine-poly-L-lysine, poly(L-phenylalanine, L-glutamicacid)-poly-DL-alanine-poly-L-lysine, and poly(L-tyrosine, L-glutamicacid)-poly-DL-alanine-poly-L-lysine, copolymers of L-arginine withtryptophan, tyrosine, or serine, copolymers of D-glutamic acid withD-lysine, copolymers of L-glutamic acid with lysine, ornithine, ormixtures of lysine and ornithine, and poly (L-glutamic acid), and theuse of amines and other positively charged amino acids bound topolymeric configurations to mimic the cationic polymeric activities sodescribed.

Examples of hydrophobic co-polymer blocks include, but are not limitedto, alkanes, alkenes, alkynes, poly(isobutylene), polyesters such aspoly(caprolactone) (PCL), poly(lactic acid) (PLA), poly(glycolic acid)(PGA), and copolymers therefrom (PLGA), polyamides such as nylon(6,6)and Nylon(12), polyurethanes, poly(propylene oxide), poly(tetramethyleneoxide), polyethylene, polypropylene, polystyrene, poly(acrylates) suchas polymethyl acrylate (PMA), poly(methacrylates) such aspoly(methylmethacrylate) (PMMA), poly(sulfones), poly(etheretherketones)(PEEKs), poly(phosphazines), poly(carbonates), poly(acetals) andpoly(siloxanes).

As described supra, a host of additional adsorbing agents can beutilized in the polymer construction.

As will be understood by the skilled artisan upon reading thisdisclosure, triblock configurations can be used. An exemplary blockco-polymer comprising a triblock configuration is PLURONIC® F127, alsoreferred to as Poloxamer 407, containing a poly(ethylene oxide)hydrophilic block (“PEO”), a poly(propylene oxide) hydrophobic block(“PPO”) and another PEO block. Other block co-polymers for use in thepresent invention may contain only one hydrophilic block and onehydrophobic block, or may contain several alternating blocks, forexample the PPO-PEO-PPO block co-polymers (PLURONIC®, block co-polymersbased on ethylene oxide and propylene oxide, BASF, Florham Park, N.J.).Additional exemplary PLURONIC block co-polymers useful in the presentinvention include, but are not limited to, PLURONIC 10R5, PLURONIC 17R2,PLURONIC 17R4, PLURONIC 25R2, PLURONIC 25R4, PLURONIC 31 R1, PLURONIC F108 Cast Solid Surfacta, PLURONIC F 108 Pastille, PLURONIC F 108 Prill,PLURONIC F 108NF Prill Polaxamer 338, PLURONIC F 127 Prill, PLURONIC F127 NF, PLURONIC F 127 NF 500 BHT Prill, PLURONIC F 127 NF PrillPoloxamer 407, PLURONIC F 38, PLURONIC F 38 Pastille, PLURONIC F 68,PLURONIC F 68 Pastille, PLURONIC F 68 LF Pastille, PLURONIC F 68 NFPrill Poloxamer 188, PLURONIC F 68 Prill, PLURONIC F 77, PLURONIC F 77Micropastille, PLURONIC F 87, PLURONIC F 87 NF Prill Poloxamer 237,PLURONIC F 87 Prill, PLURONIC F 88 Pastille, PLURONIC F 88 Prill,PLURONIC F 98, PLURONIC F 98 Prill, PLURONIC L 10, PLURONIC L 101,PLURONIC L121, PLURONIC L 31, PLURONIC L 35, PLURONIC L 43, PLURONIC L44, PLURONIC L 44 NF Polaxamer 124, PLURONIC L 61, PLURONIC L 62,PLURONIC L 62 LF, PLURONIC L 62D, PLURONIC L 64, PLURONIC L 81, PLURONICL 92, PLURONIC L44 NF INH surfactant Polaxamer 124, PLURONIC N 3,PLURONIC P 103, PLURONIC P 104, PLURONIC P 105, PLURONIC P 123Surfactant, PLURONIC P 65, PLURONIC P 84, and PLURONIC P85. Whereapplicable, all particle sizes of the block co-polymers are included,for example PLURONIC F127 and PLURONIC F87 are available as prill andmicroprill products. Non-ionic surfactants, for example, containing ahydrophobic segment and a PEO block are considered here as blockco-polymers. The use of these commercially available agents specificallyin combination in formulations with bifunctional copolymers heredescribed including cationic and other adsorbing moieties coupled withhydrophilic or non reactive elements is identified.

Additional exemplary block or graft co-polymers or dendrimers which canbe used in the present invention are disclosed in U.S. Pat. No.5,578,442 and U.S. Pat. No. 5,834,556, as well as patents: U.S. Pat.Nos. 462,990; 5,627,233; 5,567,440; 5,849,839; 5,820,882; 5,380,536;5,232,984; 6,231,892; 6,743,521; 7,316,845; 2,286,590; 6,596,267;7,029,688; 6,350,527; 6,652,902. Teachings of each of which are hereinincorporated by reference in their entirety.

By “formulations” it is meant the particular mixture of base chemicalsand additives required for a product.

As mentioned previously, the examples of multi-functional orbifunctional copolymers described above can be used in formulations fortopical ophthalmic use and for coating extraocular and intra-cornealdevices. However, the invention is not limited only to those entitiesdescribed above, other variations or types of graft and blockco-polymers having properties described herein is an embodiment of theinvention. The invention is not limited only to those block and graftco-polymers named and described in detail herein.

The block or graft co-polymers are included in formulations for use inthe methods of the present invention at weight percent concentrationsranging between 0.001% and 40%, more typically 0.01% to 25%, of theformulation. An effective amount is claimed. In the Examples in whichthe formulation is described at present, the amounts of co-polymers arebetween about 0.1% and 2%. In addition the amount of bi-functionalco-polymer can fall within other smaller ranges e.g., 0.01% to 3%, 0.1%to 2.5%, or 0.5 to 2%.

The combinations of different copolymers can be that each is(weight/weight) between 0.01% and 2% but can have ranges high as wide as0.01% and 5%.

An effective amount is an amount that increases the wettability of agiven biological surface, or contact lens surface; or that changes ormodifies the surface in a favorable way; such modification can includebut is not limited to steric stabilization, steric repulsion, highsurface water retaining capacity, charge neutrality, surface exclusioneffects, or osmotic repulsion. The extent of increase in wettabilitywill vary with the application and with the disease or disease symptomthat is being treated or prevented. An effective amount is an amountthat leads to beneficial biological surface modificationcharacteristics, as described above.

The bi-functional copolymers are used in any of these concentrations toimpart favorable characteristics to a product for use in humans or othermammals either with or without an active pharmaceutical ingredient. Anactive pharmaceutical ingredient can be added to these formulations andthe active agent will be used in weight percent concentrations frombetween 0.0001% to 40%, more typically from 0.01% to 10%. The copolymerscan be used with a preservatives such as, but are not limited to,Polixetonium, polyquaternium-42, Polyquaternium-1, Polyquat,Alkyl-hydroxy benzoate preservatives, parabens, hydrogen peroxide,benzalkonium chloride, cetylpyidimine chloride, cetalkonium chloride,sodium perborate, Purite, disappearing preservatives, Polyhexamethylenebiguanide (PHMB), chlorobutanol, Benzododecinium bromide, “Ionicbuffered system”, povidone, silver, silver sulfate, betadine, and otherantiseptics and proprietary and non-proprietary preservatives. Thepolymers herein described can be used in combination with surfactants oremulsifier. Surfactent, means a chemical agent capable of reducing thesurface tension of a liquid in which it is dissolved. Emulsifier means asubstance which stabilizes an emulsion by increasing its kineticstability.

An important aspect of this invention is formulations combiningbi-functional copolymers with one or more preservatives, surfactants,and/or emulsifiers.

Polydispersity in these graft and block co-polymers can be eitherminimal or there can be significant amounts of polydispersity, and theformulations can still be effective. Viscosity can important in theseformulations. Typically, with current technology, higher viscosity eyedrop formulations lead to longer surface changes on an eye but areassociated with visual blur and dissatisfaction by the user. There areadvantages to lower viscosity agents, and in certain formulationsdescribed herein a lower viscosity can be beneficial as there will beless visual blur imparted when an eye drop is applied. In the currentembodiment, the viscosity is approximately 2.7 centipoise (cP), however,it can be higher or lower. 2.7 cP is lower than the viscosity of mostcommercially available artificial tears. In some embodiments a higherviscosity formulation may be preferred, however. Viscosity can beadjusted by adjusting concentrations of the graft and/or blockco-polymers, or by adjusting other excipients to affect viscosity.Useful viscosities are in the range of 1 cP up to and including 100 cPor higher, but typically the viscosity would be between 2 and 30 cP.

Additional exemplary components which can also be incorporated intopharmaceutical formulations and coatings for use in the presentinvention include, but are not limited to PLURONIC gelling agents suchas, but not limited to F127, F108 as well as additional PLURONIC agentslisted supra. Furthermore, in one embodiment, these components are usedat fractions below that required for gelling activity.

Other components (either active or inactive ingredients) which can beincluded in these pharmaceutical formulations include, but are notlimited to, lipids, oils, surfactants, water, lubricating polymers,typical surfactants, buffers, salts, physiologic ions, proteins, topicalemollients, excipients typically used in oral, topical, mucosal,dermatologic and ophthalmic formulations, lubricants such as PEG 400,carboxymethylcellose, hydroxypropyl methylcellulose, mineral oil,propylene glycol, glycerin, hypromellose, white petrolatum, polyvinylalcohol, liposomes, mannitol, hydroxypropyl guar, dextran 70,viscoelastics, guar gum, alginates, and hyaluronic acid, as well ascombinations thereof. Specifically, the use of the herein describedcopolymers for ophthalmic indications in combination with active agentsdescribed in the OTC Monograph 21CFR349.14 is claimed. Additionalingredients may include those routinely included in shampoos, soaps, andconditioners. Such components may be included in the formulations invarying percentages ranging from less than 0.1% to 99% w/w %, morepreferably less than 1% to 10% w/w %. These are wt %'s relative to thetotal formulation.

Other components which can be included in these pharmaceuticalformulations include preservatives such as, but are not limited to,Polixetonium, polyquaternium-42, Polyquaternium-1, Polyquat,Alkyl-hydroxy benzoate preservatives, parabens, hydrogen peroxide,benzalkonium chloride, cetylpyidimine chloride, cetalkonium chloride,sodium perborate, Purite, disappearing preservatives, Polyhexamethylenebiguanide (PHMB), chlorobutanol, Benzododecinium bromide, “Ionicbuffered system”, povidone, silver, silver sulfate, betadine, and otherantiseptics and proprietary and non-proprietary preservatives. Also,PLL-g-PEG, or other cationic components of graft or block co-polymersmay act as a preservative. Included in this invention, also, is the useof cationic moieties in the graft or block co-polymers to serve as apreservative for a formulation containing these agents. Poly (L) lysine,alone, not coupled with a second polymer, as an ophthalmic preservative,for example, is included as an invention in this filing.

Further, in some embodiments, the formulations and coatings may includeone or more additional active pharmaceutical ingredients. Examplesinclude, but are in no way limited to anesthetics, antibiotics,anti-virals, anti-inflammatory agents, intraocular pressure loweringagents, artificial tears, lubricating products, dilating agents,immunosuppressives, anti-angiogenic agents, proteins, peptides,neuroprotectants, small molecules, growth factors and antibodies. Byactive pharmaceutical ingredient or API it is also meant to be inclusiveof genes and/or gene transfer agents such as adenovirus, AAD and nonviral vectors for which transfer and transfection is enhanced viaformulations of the present invention. Formulations may be used inaccordance with the present invention to deliver an activepharmaceutical ingredient which acts locally at the surface or in thetissue to which it is delivered. Alternatively, or in addition,formulations may be used in accordance with the present invention todeliver an active pharmaceutical ingredient which is then absorbedthrough the tissue and has a systemic or distal effect. In someembodiments, delivery of the active pharmaceutical ingredient isfacilitated by external energy such as, but not limited to,iontophoresis, sonic energy, heat, microneedles, micropore creatingdevices such as lasers or high pressure gas.

Pharmaceutical formulations are a composition suitable for internal,topical, or ocular administration to an animal, including humans. Thecarrier is a pharmaceutically acceptable excipient in which theco-polymer is admixed.

“Eye lubricant” can be defined as the API included in artificial tearproducts based on the FDA OTC monograph 21 CFR 349.14 for ophthalmics.

pH of formulations of the present invention is in a physiologic rangedepending upon the site of administration and the site of the biologicalsurface or membrane that is to be modified. Typically, the pH is above3, e.g., above 5.6 and below 9. The pH is preferably between 6.5 and 8.Hypertonic and hypotonic formulations are claimed.

The ability of a pharmaceutical formulation comprising a graft or blockco-polymer with a positively charged or hydrophobic moiety and ahydrophilic moiety to change wettability and adhere to and lubricate andmodify a biological surface was demonstrated. Thus, the triboloogicalproperties of the surface was modified. Importantly, these formulationsmay have very low viscosity (lower than most artificial tear products onthe market). For example, the formulation tested in humans had aviscosity of 2.7 cP. Thus, the results are not due to a simplemechanical thickening of the tear film.

The ability of formulations comprising either the graft co-polymerPLL-g-PEG or the block co-polymer F127 to change wettability wasdemonstrated in vitro via contact angle, experiments, as described inExample 2. FIG. 1 and FIG. 2 demonstrate the ability of exemplaryformulations comprising either 1, 0.5 or 0.1% weight/weight PEG-PLL or1% F87 or F127 weight/weight, respectively, to decrease the contactangle of water as compared to a control of deionized water. The abilityof these formulations to decrease contact angle is indicative of theirability to adhere to negatively charged surfaces such as biologicalsurfaces and/or membranes and change wettability of the biologicalsurface and/or membrane. Through the understanding of one skilled in theart, these formulations will thus also impart surface modifications suchas steric stabilization on the tissue to which the bi-functional copolymers adhere.

As described in Example 3, Ex vivo experiments in porcine eyes alsoshowed a delay in pre-corneal water evaporation rates of approximately 2seconds longer compared to a control of Systane® lubricant eye drops(Active Ingredients: Polyethylene glycol 400 0.4% and propylene glycol0.3%. Inactive Ingredients: boric acid, calcium chloride, hydroxypropylguar, magnesium chloride, potassium chloride, purified water, sodiumchloride, zinc chloride.

As described in Example 1, experiments in a rabbit model also showedlack of toxicity, in other words, safety, for a formulation comprising1% glycerin with 1% PLL-g-PEG and 1% F127 as delivery of suchformulation of the present invention to a sensitive mucosal surfaceshowed no visible irritation in the Draize test and no histopathologicalchanges (14 day histopathology study).

Furthermore, as described in Examples 4 and 5, performance of theseblock and graft copolymer formulations was assessed in the eyes of humanvolunteers. Initial informal tolerability studies showed the eye dropwas well tolerated. For example, of the multiple initial humanexposures, there was no irritation or discomfort seen or reported in anysubject. Additionally, there were no reports of blur followinginstillation of 50 microliters or less. Actually, the eye drop wasdescribed as soothing on several occasions. As a further safety step,one volunteer received 1 milliliter of solution in the eye, multipletimes, and there was no irritation or discomfort. The ability to treatdry eye was then assessed using tear film break up time (TFBUT). TFBUTis indicative of the rate of evaporation of tears from the surface ofthe eye. The longer the TFBUT, the wetter the eye between blinks and theless likely a patient will have dry eye signs and symptoms. The firstassessments were informal and performed in only a couple of subjects,then a larger masked, randomized controlled study was carried out.Administration of a formulation comprising 1% glycerin with 1% PLL-g-PEGand 1% F127 resulted in prolongation of TFBUT. In the first informalstudies, the TFBUT was measured using fluorescein staining. The TFBUTwas prolonged versus active comparator on one occasion at early timepoints, and that increase was maintained for three hours. Subjectiveobservation of the tear film break up into smaller patches was confirmedvia a non-invasive wavescan of the tear surface. The larger, randomizedcontrolled trial studied three groups of patients (none/mild dry eye,mild/moderate dry eye, and moderate/severe dry eye). The total number ofsubjects was 18. Sixteen were analyzed with a noninvasive TFBUTassessment tool called the Tearscope™. In this study, there wasstatistically significant prolongation of the TFBUT at the 15 minutetime point and the two hour time point versus active comparator (aleading OTC dry eye artificial tear). The two additional subjectsunderwent wavescan assessments as an alternative to the Tearscope Plus™the results in these two subjects also showed prolongation of the tearfilm compared to control. Accordingly, one embodiment of the presentinvention relates to use of these pharmaceutical formulations aslubricating opthalmic eye drops for treatment of, for example, dry eyesyndrome and contact lens intolerance. As will be understood by theskilled artisan upon reading this disclosure, however, alternativeophthalmic delivery means including, but not limited to, intraocular,periocular, conjunctival, subconjunctival, transconjunctival,peribulbar, retrobulbar, subtenons, transscleral, topical gel, topicaldispersion, intraorbital, intrascleral, intravitreal, subretinal,transretinal, choroidal, uveal, intracameral, transcorneal,intracorneal, intralenticular (including phakia and psuedophakia), andin or adjacent to the optic nerve, can be used. The invention can beused with polymeric and other devices for prolonged ophthalmic drugdelivery. The invention can be used with depot formulation to ease thetolerance of the eye to prolonged drug exposure.

Dry eye syndrome is a common and irritating problem for approximately 60million Americans. Five million Americans suffer from an advanced formof this condition. Dry eye syndrome can be due to a deficiency of theaqueous, lipid, or mucin component of tears. Dry eye syndrome is alsorelated to inflammation on the surface of the eye and irregularities ofthe tear film components. Aging and hormonal changes can play a role.Environment, as well as contact lenses and refractive surgery, caninitiate and/or exacerbate the problem.

Dry eye is treated with eye drops, punctual occlusion, and occasionallysystemically administered medicine, although eye drops are the mainstayof treatment. While most eye drop treatments are availableover-the-counter, there are two prescription eye drops inclusive ofsteroids and cyclosporine that help with dry eye. Most over-the-counterdry eye syndrome products are washed out through the lacrimal drainagesystem quite quickly thus rendering relief minimal. For example, in atleast two studies by Alcon, Systane had a beneficial effect on tear filmbreak up time that lasted up to 30 minutes. These studies demonstrateSystane™ is one of the longest acting over-the-counter dry eye products.The PLL-g-PEG formulation shows results improving tear film break uptime beyond two hours.

Accordingly, there is need for formulations of the present invention fortreatment of this condition.

Formulations can also be used in accordance with the present inventionto provide for prolonged coating of the ocular surface via themultifunctional properties of the graft and/or block co-polymers.Without being bound to any particular theory, it is believed that thehydrophilic domain(s) of the block or graft co-polymer, which keep theeye lubricated and help to retain tears on the surface, are anchored atthe surface of the eye (cornea and conjunctiva) for an extended periodof time through primarily interaction of positively charged domain(s) onthe block, graft, or backbone of the graft copolymer and secondarilythrough hydrophobic domains on the block, graft or backbone of thepolymer with the surface of the eye through either electrostaticattraction with the negatively charged areas on the surface of the eyeor hydrophobic interactions with the hydrophobic regions of the eye thatplay a role in dry eye syndrome. Similarly, these polymers couldinteract with natural mucins and be effective in adhering to mucinsand/or keeping soluble mucins in the tear longer, and/or increasing theeffectiveness and natural protections provided by mucins. Theformulations and use of the graft and block copolymers described hereinmay also have beneficial effects based on the steric stabilization ofbiological surfaces. The protection provided may help break the viciouscycle of inflammation, cellular injury, and discomfort.

Formulations comprising a bi-functional co-polymer and an anestheticagent such as proparacaine can be used in accordance with the presentinvention to prolong the anesthetic effect and/or reduce the acutecorneal surface changes seen commonly in the clinic and operating roomassociated with decreased sensation of the cornea and a decreased blinkrate. Such formulations can be applied prior to examination and/orpre-operatively, and will help maintain a more normal corneal epithelialsurface. Furthermore, use of the formulation will help reduce bacterialadherence at sites of any procedures performed on the eye and will helpreduce post-procedural infection.

Formulations described herein also provide a safe, nonirritatingexcipient for ophthamological formulations with one or more additionalactive pharmaceutical ingredients. For example, one aspect of theinvention is the use of the block and graft copolymers described hereincombined with active agents for treating ophthalmic disease. Theirinclusion may reduce the irritation or tachyphylaxis (associated withsome eye drops), or simply provide an additional lubricating andwettable ocular surface enhancing the comfort and acceptance of eyedrops. Exemplary additional active pharmaceutical ingredients forophthamological uses include, but are not limited to, lubricants anddemulcents, as described supra, antibiotics (fluoroquinolones,vancomycin, cephalosporin, gentamycin, erythromycin, azithromycin, sulfadrugs, bacitracin, gatifloxacin, levofloxin, moxifloxacin, ofoxacin),acetazolamide, antazoline, aspirin, atropine, azelastine, bacitracin,betaxolol, bimatoprost, botanical drugs including zeaxanthine lutein,lycopene brimonodine, brinzolamide, carbachol, carteolol, ciprofloxacin,ofloxacin, cromalyn, cyclosporine, cyclosporine pro-drugs andcyclosporine derivatives, other immunomodulators, dapiprazole,dexamethasone, diclofenac, dipivifren, dorzolamide, epinephrine,erythromycin, fluoromethalone, flurbiprofen, gentamycin, glaucomamedications (prostaglandins, carbonic anhydrase inhibitors, epinephrineor alpha-agonists, beta-blockers), gramicidin, homatropine,hydrocortisone, hyoscine, keterolac, ibuprofen, ketotifen, latanaprost,levobunolol, levocabastine, levofloxin, lotepprednol, medrysone,methazolamide, metipranolol, naphazoline, natamycin, nedocromil,neomycin, neuroprotective agents, nonsteroidal anti-inflammatory agents,nepafanec, norfloxacin, ofloxacin olopatadine, oxymetazoline,pemirolast, pheniramine, phenylephrine, pilocarpine, povidone,prednisolone, proparacaine, scopolamine, tetracaine, steroids,sulfacetamide, tetrahydrozoline, hypertonic tears, timolal, tobramycin,travaprost, trifluridine, trimethiprim, tropicamide, unoprostone andzinc. Prodrugs and related compounds, as well as any new activepharmaceutical ingredients can be used with the block and graftcopolymers here described.

Formulations comprising a graft and/or block co-polymer can also be usedin accordance with the present invention with contact lens solutions,manufacturing, rewetting drops, or in pre-insertion contact lenstreatments. The graft and/or block co-polymers can be used with currentcontact lens care solution and rewetting drop ingredients, including,but not limited to, water, preservatives, NaCl and other salts andmineral ingredients, buffers, and other polymers and osmotic agents. Theuse of these graft and block co-polymers in contact lens relatedconsumer products to impart clinical benefits is claimed. Also claimedis the use of these graft and block copolymers in the manufacture,coating, finishing, and storage of contact lenses.

Formulations comprising a bi-functional co-polymer can also be used inaccordance with the present invention with a second eye drop, such as alipid or oil based eye drop, to enhance the efficacy of the second eyedrop. In one embodiment of this use, the formulation described herein isadministered first followed by administration of the second eye drop.

Additionally, these formulations with the ability to prevent or inhibitcell adhesion are useful in preventing or inhibiting the spread ofbacterial or viral infections such as bacterial (including Chlamydial,gonorrheal, Staphylococcus epidermidis, Streptococcus pyogenes,Streptococcus pneumonia, Neisseria meningitidis and Moraxella lacunata,Haemophilus and family Enterobacteriaceae, Clostridium species,gram-negative anaerobic bacilli, and Peptostreptococcus species, N.gonorrhoeae, C. trachomatis, staphylococcal species and S. pneumonia) orviral keratitis or conjunctivitis (including herpetic keratitis andadenovirus conjunctivitis) and in preventing and/or inhibitinginflammation caused by adherence of inflammatory cells and proteins tothe eye surface. The invention may be useful in treating or preventingamebic, protozoal, mycobacterial and other types of keratitis andconjunctivitis. There may be a benefit in treating sterile ulcers,ectasia, and corneal or scleral melts. The anti-inflammatory effect mayalso be due to inciting agents being kept off the corneal andconjunctival surface decreasing the inflammatory reaction. Suchformulations will be particularly useful in neonatal, daycare, pediatricor family settings wherein outbreak of one individual can lead toinfection of many others.

For the eye, the application of a formulation in accordance with thepresent invention is expected to reduce the exposure of the surface ofthe eye to bacterial antigens, such as those from Staphylococcus Aureus,and thus may provide an important part of the treatment for staphmarginal disease. There also may be benefits to patients with allergicconjunctivitis including giant papillary conjunctivitis (limiting directtissue exposure to antigens), and for patients who have had previousfiltering surgery with thin ischemic blebs, as therapy with aformulation in accordance with the present invention is expected toreduce the risk of bleb related endophthalmitis. Additionally, after acorneal abrasion application of a formulation in accordance with thepresent invention can decrease the risk of corneal ulcer development,especially in the setting of injuries sustained from vegetable matterwhich predispose a subject to fungal infections. Other conditions thatcould be treated with formulations described with this invention includeconjunctivitis and keratitis due to processes such as Moorens ulcer,Terriens marginal degeneration, ligneous conjunctivitis, toxicexposures, autoimmune conjunctivitis, and phlyectenular conjunctivitis.Use in patients with keratoprostheses is claimed. The eye drops can beused in mammals, humans, or even dogs, cats, and horses.

From experiments described herein, it is expected that formulations ofthe present invention will also be useful changing wettability and/or,steric stabilization, tribological properties, lubricating and/orpreventing adherence of unwanted proteins and cells to other biologicalsurfaces including, but not limited to, skin, mucous membranes and hair.These formulations can thus also be applied in accordance with thepresent invention to epithelial tissue of the skin, urinary, orgastrointestinal tract, mucous membrane, exposed wound surfaces,including respiratory tract mucosa, oral and nasal mucosa, vaginalmucosa, and conjunctival surfaces, and surgical and traumatic wounds andulcerations. These formulations can serve to protect skin and otherorgans from foreign protein, viral, and bacterial adherence. Benefitscan include reduction in rate of infection. Without being limited to anyparticular mechanism, it is believed that a formulation comprising agraft polymer as described above, such as PLL-PEG, adheres to thebiological surface via the PLL moiety while the PEG moiety preventsand/or reduces the adherence of potentially harmful particles. Decreasedadherence can be beneficial by decreasing pathogen or toxin exposure,thereby decreasing morbidity and mortality associated with these agents.For example, lower bacterial loads can decrease the severity ofsubsequent bacterial infection and/or exotoxin exposure, and allow thehost defenses and antibiotics better opportunity to work. Decreasingviral exposure can reduce transmission rates and possibly the severityof viral infection. These formulations are useful as well againstfungals and exposure to reactive proteins.

Accordingly, the formulations described herein have a wide variety ofuses in accordance with the present invention.

For example, these formulations can be used in accordance with presentinvention in military applications to reduce soldiers' and citizens'morbidity associated with biological or biochemical warfare attacks.

These formulations can also be used in accordance with the presentinvention to reduce rates of methicillin-resistant Staphylococcus aureus(“MRSA”) adherence in hospitalized patients, children, and the generalpopulation. MRSA is a growing concern in the health care setting.Reducing the adherence to the epithelial surface thereby reducing MRSAspread and severity and frequency of MRSA infections is expected toprove very beneficial. Such use is expandable to school situations,prehospital admission, nursing homes and chronic care facilities aswell. It is not expected that MSRA, or other pathogens, would developresistance to formulations of the invention.

These formulations can also be used in accordance with the presentinvention to reduce transmission of the common cold and influenza virusvia nasal and inhalational applications in settings including, but in noway limited to, airplanes, preschools and schools, homes of affectedviral individuals, as well as nursing homes and chronic care facilities.The block or graft co-polymers can be included in nasal sprays andproducts formulated for inhalational delivery, as is known to thoseskilled in the art. For example, a nasal spray would include the blockor graft co-polymers and benzalkonium chloride, dextrose anhydrose,edentate disodium, microcrystalline cellulose and carboxymethylcellulosesodium, polysorbate 80, and purified water. An aerosol inhaler wouldinclude the block or graft co-polymers as well as ethanol and propellantsuch as propellant HFA-134a (1,1,1,2-tetrafluoroethane).

Important utilities of the formulations exist for children and adultswith infirmities, such as immunodeficiencies, chronic disease and otherchronic conditions such as cystic fibrosis, which leave the host moresusceptible to routine illness.

Application of a formulation in accordance with the present invention toa wound just after injury or surgery, including, but not limited tocorneal abrasions, corneal surgery such as LASIK, PRK, or otherrefractive or vision correction procedures, or intraocular surgery wherean incision is made in the eye through sclera or conjunctiva/sclera,such as for cataract or glaucoma surgery. The formulation can be usedagain after cleansing, can reduce bacterial infection, and infectiouswound complications. Such formulations may be particularly useful in thefield in military medical operations, after an initial irrigation.

For these uses in humans, the formulation may be in the form of alotion, gel, liquid, spray, rinse, dissolvable wafer, or glycerin bar towhich water is added to solubilize the graft co-polymer or blockco-polymer to make it more amenable to application. Formulations can beprovided as individual or single use products or in volumes forindustrial use and/or multiple use dispensers. In addition to thebi-functional co-polymer, such formulations may comprise any and alltypical binders, excipients, and components found in cosmetic sprays,lotions, soaps, shampoos, cleansers, and oral, nasal, vaginal, and eyecare products.

Formulations of the current invention can be delivered by eye dropbottles with a nozzle on the end (volume generally between 0.2 cc and 50mL). The container may be single use disposable containing a volume ofapproximately 0.3 mL or a multi use vial containing a volume between 0.3mL and 50 MI. The material would be high density polyethylene orsimilar, and the container could be up to 500 mL for multipurposecontact lens solutions, and would have a nozzle. The solution could bestored in contact lens containers, as well.

By “eye care solution container” it is meant a container for eye caresolutions that is made of high density polyethylene or similarthermoplastic or petroleum based plastics for health care use. The eyedrop bottle is made from a mold or by blow, fill and seal methodology.The eye drop solution container has a nozzle for releasing smallerquantities of the solution than the total volume of the container. Eyecare solution container may be single use or multi-use. The volumes foran eye care solution container range from 0.1 cc to 500 cc.

Formulations comprising a bi-functional co-polymer with a positivelycharged or hydrophobic moiety and a hydrophilic moiety can also be usedin accordance with the present invention in the treatment of dry mouthin subjects with, for example, but not limited to, Sjogrens syndrome, ordry mouth during or post chemotherapy. Such formulations may furthercomprise traditional lubricants for such dry mouth conditions, such as,but not limited to, xylitol, and glycerin MouthKote.

The ability of these formulations to lubricate mucous membranes withoutirritation makes these formulations useful as sexual lubricants as well.Reduced transmission of HIV and other sexually transmitted diseases whena formulation is applied prophylactically on skin and mucosal surfacesin accordance with the present invention prior to exposure may be anadditional, if not a primary benefit in this application.

Formulations can also be used in accordance with the present inventionto protect gums in, for example, periodontal disease by reducinginflammatory cell and bacterial cell adherence to the gums. In thisembodiment, these formulations may be incorporated into, for example,but not limited to, a toothpaste or mouthwash and may be combined withflavorant and/or fluoride. Formulations which reduce bacterial adherencein the mouth, and in particular the posterior tongue and throat, arealso expected to be useful in treating bad breath or halitosis.

Formulations can also be used in accordance with the present inventionas a preoperative shampoo to reduce bacterial adherence to hair and/orskin before surgery. Since these formulations have been found to be safefor administration to the eye, they provide a useful preoperativeshampoo to remove any bacteria adhering to the eyelashes prior to eyesurgery.

The ability to reduce or prevent bacterial adherence to biologicalsurfaces such as skin and hair is expected to make these formulationsuseful in the treatment of chronic blepharitis as well, with or withoutconcomitant steroids or antibiotics. Formulations described herein maybe used on the eyelids and eye lashes, for treatment of medicalconditions and for prophylaxis. For these embodiments, the formulationmay be combined with an antiseptic or an antibiotic.

Further, the ability to reduce bacterial adherence to skin makes theseformulations useful in deodorants to reduce body odor caused by bacteriathat break down sweat.

Formulations can also be used in accordance with the present inventionon animals, including household pets, to decrease bacterial adherence totheir skin, oral cavity, or coats. In this embodiment, the formulationcan be in the form of a spray, shampoo, lotion, or beverage.

Other uses for these formulations will become evident to those skilledin the art upon reading this disclosure and are such uses areencompassed by the present invention.

The present invention also provides coatings comprising a graftco-polymer with a positively charged or hydrophobic moiety and ahydrophilic moiety or a block co-polymer with a positively charged orhydrophobic moiety and a hydrophilic moiety for extraocular devices suchas contact lenses, glaucoma stents and valves, scleral buckling hardwareand external drug delivery devices. External drug delivery devices caninclude biodegradable devices as well as permanent products such asthose made of silicon and other non bioerodible polymers. There is apotential use for intraocular devices including, but not limited to,intraocular lenses, intraocular contact lenses, glaucoma stents, tubesand valves, intracorneal implants that can be used for the correction ofpresbyopia, intraocular pressure sensors (including devices forcontinuous or microchip measurements with or without telemetry forintraocular pressure and intracranial pressure monitoring), andmicrochips used for retinal vision enhancement. Such coatings improvewearability and biocompatibility of these extraocular devices (such ascontact lenses) and decrease proteinaceous, bacterial, fungal, andparticulate adhesions to these extra-ocular devices. These coatings alsoimprove optical performance of the lenses. Coating intraocular andextraocular devices in accordance with the present invention alsoreduces epithelial adhesion and fibrosis thereto.

For example, contact lenses are often difficult to tolerate due to thediscomfort caused by the adhesion of proteins such as lysozyme anddenuded protein products and other particles to the negative charges ofthe lens surface. Other particles adhering to the lens surface includebacteria and bacterial proteins, endotoxins, antibodies, calcium, andlipids. Other uncharged particles, and even negatively charged particlescan build up on the lens surface once this cascade of protein adhesion,cellular and/or particulate adhesions to the lens is initiated. Wearingof contact lenses with these materials adhered to the contact lens canlead to discomfort, allergic reactions, ocular irritation, foreign bodysensation, epithelial breakdown and dysfunction, and bacterialkeratitis.

By binding the surface of a contact lens with positively charged and/orhydrophobic domains of block or graft co-polymers included in topicalformulations of the present invention, the aggregation of proteindeposits during wear time on the lens is and will be reduced and contactlenses are and will be better tolerated. The lens surface is also morelubricious, and the steric stabilization as well as high surface waterretaining capacity and charge neutrality can impart numerous benefits inthis clinical setting. With protection provided by the polymers hereindescribed, the contact lenses may also be more wettable and feel morecomfortable with the concomitant use of formulations described herein aswell as other ophthalmic products. Formulations can be provided inaccordance with the present invention as eye drops to improve contactlens wearability, as a finishing rinse after lens manufacture, instorage solutions, in lens cleaning solutions, in rewetting drops, as adrop to be used prior to placing the lens in the eye, and/or as anactive preservative in contact lens preparation and storage. Theseformulations thus may be used in accordance with the present inventionin the form of a one time application or repeated application. Contactlenses and intra- and extra-ocular devices can be made morebiocompatible with the coatings of the graft and block co-polymers asdescribed herein.

There are well known similarities, and there exists an establishedclinical relationship between dry eye syndrome and contact lensintolerance. In studies described in examples 4 and 5 in this invention,it has been shown clinically that the formulations and uses of graft andblock copolymers described herein conferred meaningful and measurablebenefit in dry eye subjects. These results are highly suggestive of asignificant opportunity for clinical benefit in the contact lens use andcontact lens intolerance clinical setting. By way of interpolation, itfollows that there will be similar benefits conferred in the use ofthese graft and block copolymers on other extraocular- and intra-cornealdevices.

It can be foreseen that some intra-corneal or extra-ocular devices willhave prolonged biological use (for example extended wear contact lens,and surgically implanted devices). Under conditions of high stress (suchas ionic, salts or other electrolyte media, enzymatic, pH, ortemperature) and likely prolonged time in aqueous electrolyte media, theelectrostatic or hydrophobic interactions between the graft and blockcopolymers and the intra- or extra ocular devices may be reduced to somedegree. Additionally, the tear film, sub-epithelial, or intraocularfluid will likely not (unless a specific intervention has beenundertaken for that purpose) be a reservoir for replacement graftcopolymers as they naturally degrade over time. The benefit conferred bythe polymers to the device may, therefore, wear off over time. Thus,there may be advantages to covalently immobilizing the block or graftcopolymers herein described to the intra or extra ocular device duringmanufacturing or finishing. It follows, therefore, that covalentlybonding, embedding during manufacture, or covalently immobilizing thesegraft and block co polymers to the devices may be beneficial. It hasbeen shown that covalent immobilization does not necessarily result inless efficient packing of a graft co-polymer (Blattler, et al. Langmuir2006, 22, 5760-5769). An aldehyde plasma polymer interlayer withreductive amination can be used in such a setting. Other methods forcovalently immobilizing the block or graft copolymers can alsoaccomplish the same result of developing a more stable and resistantcoating on the surface of the device.

Use of block or graft co-polymer containing formulations to coatintracorneal devices also reduces particulate, cellular, epithelial,and/or fibrocyte adhesion. Intracorneal inlays, may benefit from the useof these graft and block copolymers. Reductions in the risk of infectionand clotting or blockage by protein or fibrin debris, cellularproliferation, inflammation suppression, and steric repulsion are allways in which these polymers may confer advantages to extra- orintraocular devices, including intracranial implants.

The descriptions provided above are not to

be construed as limitations of the present invention and its variousembodiments described herein, but rather, are described to fully allowone skilled in the art to make and use the invention along withmodifications, adaptations, and alterations, all being within the spiritand broad scope of the present invention as fully defined herein.Similarly, the following nonlimiting examples are provided to furtherillustrate the present invention. Percentages are weight percents.

EXAMPLES Example 1

The following topical formulation was made and tested:

1% F127

1% PLL-g-PEG

1% glycerin

sterile water for injection

sodium phosphate buffer

mannitol to adjust osmolality;

pH between 6.5 and 7.5 and osmolality 274-350 mOsm/kg;

Sterile Filtration

This formulation was tested in the Draize test and no ocular irritationwas observed.

This formulation was also tested in two volunteer humans and noirritation was reported. Tear film break up time in one volunteer wasextended compared to the control eye for over two hours afterinstillation of the eye drops. On close inspection under microscopy,after instillation, the tear film of the eye treated with the PLL-g-PEGformulation displayed a more stable tear film. This stability waspresent at greater than three hours after eye drop instillation.Furthermore on analysis using a Tearscope to evaluate the tear film,compared to an eye that received a commercially available product, theeye treated with the PLL-g-PEG polymer showed an increase in tear filmbreak up time by 30 seconds compared to control, on one occasion, at onehour after instillation of the eye drops.

This formulation was also applied to a contact lens and tested. One dropwas placed on each side of a contact lens. The lens was placed in ahuman eye and the patient wore the contact lens comfortably for 8 hours.The treated lens was described as being more comfortable in the eye thanan untreated control lens. This formulation was also mixed 1:1 with acommercially available preserved multi-purpose contact lens solution andthe lens was stored in the solution overnight. The lens was worn thenext day comfortably. Again, the treated lens was more comfortable thanan untreated control lens. One embodiment of the copolymers hereindescribed has thus been used as an eye drop, used in combination withother polymers, used with contact lenses, and formulated with apreservative. The subject felt decreased awareness of the contact lensand less resistance and discomfort when blinking.

Example 2

An in vitro assessment was made of the ability of 1%, 0.5%. and 0.1%PLL-g-PEG formulations and 1% formulation of F127 and F87 to decreasecontact angle of water on a polystyrene surface. The contact angle of adrop of these had a much lower contact angle on a freshly cleanedpolystyrene surface compared to a drop of deionized water. Thisexperiment demonstrates the ability of these formulations to changewettability and adherence to negatively charged surfaces such as the eyeand epithelium.

Example 3

An ex vivo assessment in porcine eyes was made of the ability of aPLL-g-PEG formulation of the present invention to decrease evaporation.Porcine eyes were obtained fresh and refrigerated. They were stored inan oil bath, which caused changes to the epithelial surface and alsopartially denuded the epithelial surface allowing for faster evaporationof an aqueous solution. The evaporation rates of small areas weremeasured and Systane was applied with no change in evaporation rateafter a single rinse. A single rinse with the PLL-g-PEG formulationshowed a reduction in evaporation time by a mean of two seconds. Fiveeyes were tested. The formulation was: 1% F127, 1% PLL-g-PEG, 1%glycerin, sterile water for injection, sodium phosphate buffer, mannitolto adjust osmolality.

Example 4

Two tests were performed in human eyes with a PLL-g-PEG formulation ofthe present invention.

The formulation was: 1% F127, 1% PLL-g-PEG, 1% glycerin, sterile waterfor injection, sodium phosphate buffer, mannitol to adjust osmolality.

The first involved the time a human eye could be held open before theneed to blink based on discomfort. The longer an eye could be held openwithout pain implies a better tear film covering over time. In thisexperiment, one eye received a single eye drop of Optive™ while theother eye received a single eye drop of the PLL-g-PEG formulation eyedrop. The control eye could be held open 30 seconds after instillationwithout discomfort repeatedly at most time points (every 10 to 15minutes) from t=5 minutes to t=120 minutes. The eye treated with thePLL-g-PEG formulation could be held open without discomfort for timesranging from 40 to 60 seconds at most time points.

The tear film was also assessed by Fluorescein tear film break up time.At t=200 minutes after instillation, there was a longer time to TFBUT byat least 5 seconds in the PLL-g-PEG treated eye. Additionally, the breakup occurred with a much smaller area of change on the surface of theeye. These results were confirmed subjectively with the wavescan devicelooking at tear film reflectance of light and aberrations.

Example 5

The formulation was: 1% F127, 1% PLL-g-PEG, 1% glycerin, sterile waterfor injection, sodium phosphate buffer, mannitol to adjust osmolality.

A randomized, masked, active comparator controlled study was carried outin eighteen subjects. The study endpoints included subjective responseto the eye drops and the tear film break up time compared topreinstillation values at t=15, 30, 60, and 120 minutes. Noninvasivetear film break up time (NIBUT) was performed using the Tearscope™ in 16subjects; fluorescein break up time (FBUT) at 120 minutes was alsomeasured in these sixteen subjects. Two subjects were tested withwavescan techniques to look at tear film break up time with that newtechnology. A questionnaire was administered to subjectively assess theacceptability of the eye drops in all subjects. The principalinvestigator was not affiliated with the product and is a full professorat an academic eye institute. The principal investigator was responsiblefor the trial. IRB approval was obtained, and the trial was registeredat clinicaltrials.gov, as per FDA requirements. The comparator was aleading over-the-counter product with the following formulation: ActiveIngredients: Polyethylene glycol 400 0.4% and propylene glycol 0.3%.Inactive Ingredients: boric acid, calcium chloride, hydroxypropyl guar,magnesium chloride, potassium chloride, purified water, sodium chloride,and zinc chloride. In the Tearscope portion with sixteen subjects, threegroups were studied: the first group of five subjects had none,occasional, or mild dry eye symptoms. The second group of five subjectshad mild to moderate dry eye symptoms. The third group of six hadmoderate to severe dry eye symptoms. RESULTS: Questionnaire: there wasno difference overall between the acceptance of the eye drops. See Table1.

TABLE 1 Preferred at 5 minutes after instillation Sample formulation 4Active comparator 5 No difference 7Vision changes: No change: 100% Eyeon; No change: 87.5% (2 said visionwas blurrier after active comparator was received. Comments at two hoursif any: Equivalent for active comparator and study formulation.Regarding slit lamp exam, there were no adverse events in any eyes, andthere were no changes in the anterior segment exam in any eyes. NIBUTresults for the entire cohort: 15 minute time point showed NIBUT timeincrease from baseline at plus 14.67 sec (p<0.05) for sampleformulation, while active comparator was 7.4 seconds longer thanbaseline (p=0.3). Thus, the new formulation essentially doubled the tearfilm break up time benefit of the active comparator at 15 minutes. FBUTwas significantly longer than active comparator at 120 minutes; superiorby 4.92 sec (p<0.12). See FIGS. 3 and 4.

The two subjects who had wavescan evaluation showed results thatsupported the conclusions in the larger study, for example, the meanTFBUT was longer for the sample formulation than active comparator atall time points.

Furthermore, a sub group analysis was performed. In the main group of 16subjects there was one subject in each group that had unexpectedly longpre-instillation TFBUT. These long initial values distorted the relativecontributions of the artificial tears to NIBUT for the other subjects.When these three outliers were removed, there was superiority inextending NIBUT for the sample formulation at every time point. The datawas evaluated by seconds over baseline, and by percentage change frombaseline. See FIGS. 5 and 6.

Thus, this randomized, controlled, masked trial supports use of theinvention for treatment, including prevention, of dry eye, contact lensintolerance, ocular irritation, and other disorders of the skin andmucous membranes.

Example 6

There is a need for compositions for treating injuries that can becustom loaded with therapeutic agents based on the suspected bacterialspecies in a wound on an external body surface (e.g., the skin).Non-restrictive loading could potentially reduce bacterial colonizationby orders of magnitude and reduce infection rates in compromisedpatients with contaminated wounds. The following results indicate thatcompositions containing a copolymer having a positively charged, orhydrophobic, or covalent bonding moiety and a hydrophilic moiety areuseful as a carrier for antibiotics. Such compositions may be usedalone, as an adjunctive therapy to wound treatment, or to protectagainst pathogen or toxin attack (e.g., from bacterial infection).

A composition containing a copolymer having a positively charged, orhydrophobic, or covalent bonding moiety and a hydrophilic moiety isformed into a wound management device or formulated in a pharmaceuticalcomposition. The copolymer composition is tested on a mouse model ofwound healing. Mice treated with the copolymer composition have fasterand/or more complete wound healing compared to untreated mice. Micetreated with the copolymer composition also have fewer or less bacterialinfection compared to untreated mice.

The copolymer composition is loaded with the antibiotic amikacin.Amikacin release from the copolymer composition is measured. The openwounds of the mice may be passively or actively infected with bacteria.Bacterial growth inhibition of P. aeruginosa is observed for woundstreated with the copolymer composition. Treatment of wounds with acopolymer composition with antibioitics results in inhibition ofbacterial growth and progress in wound healing compared to mice treatedwith the copolymer composition alone or with no treatment.

The ability to customize the antibiotic choice is advantageous becauseit allows clinicians to tailor treatment regimens based on known orsuspected bacterial species present. Copolymer compositions of theinvention allow for high release concentrations of antibiotics. Theseexperiments indicate that incorporation of antibiotics into a copolymercomposition provides a local drug delivery system that can be used aloneor in conjunction with wound treatment therapies.

Example 7

There is a need for nanoparticle compositions for treating onasal andinhalational conditions that can be custom loaded with active ortherapeutic agents based on the treatment required. Nanoparticles coatedwith a copolymer having a positively charged, or hydrophobic, orcovalent bonding moiety and a hydrophilic moiety can be used to deliveractive or therapeutic agents for inhalational applications. Forinhalation, the nanoparticle size should be 10 microns or smaller indiameter, about 1 micron or less in diameter. The copolymer coatednanoparticles are formulated with a pharmaceutically acceptable carrier.The copolymer coated nanoparticles can be used in a pharmaceuticalcomposition that is administered to the oral, nasal, or respiratorysurfaces (e.g., in the throat) using an aerosol or nasal formulation(e.g., a spray). The copolymer coated nanoparticles contain active ortherapeutic agents, including drugs (e.g., an antibiotic), menthol, orother numbing compound for treatment of coughs or sore throats. Thecopolymer coated nanoparticles deliver the active and therapeutic agentswhen contacted to oral, nasal, or respiratory surfaces (e.g., mucousmembranes). Thus, the copolymer coated nanoparticles are used to deliveractive and therapeutic agents.

We claim:
 1. A method for prolonging the stability of an ocular surfacetear film of a subject, comprising contacting said ocular surface ofsaid subject with a composition comprising a graft copolymer having apositively charged PLL backbone and one or more hydrophilic watersoluble side chains selected from the group consisting of polyethyleneglycols, polysaccharides, poly(meth)acrylates, polyacrylamides,polyvinyl alcohol (PVA), polyethyloxazoline, poly-N-vinyl pyrrolidone(NVP) and poly boronic-based polymers, in an amount and for a durationso as to prolong the stability of said tear film of said subject.
 2. Themethod of claim 1 wherein said graft copolymer creates a covalentlinkage between said graft copolymer and said ocular surface.
 3. Themethod of claim 1, wherein said graft copolymer creates an electrolyticinteraction between said graft copolymer and said ocular surface.
 4. Themethod of claim 1, where the PLL backbone is selected from the groupconsisting of: poly(L-lysine) (PLL), polylysine, and polyornithine. 5.The method of claim 1, where said polyethylene glycols are selected fromthe group consisting of polyethylene glycol, polypropylene glycol, andmixtures thereof.
 6. The method of claim 1, where said polysaccharidesare selected from the group consisting of dextran, carboxylatedcellulosics, carboxymethyl cellulose, natural gums, carageenan,alginates and hyaluronic acid and mixtures thereof.
 7. The method ofclaim 1, where poly boronic based polymers are selected from polymershaving boronic-based functional groups selected from the groupconsisting of phenylboronic acid (PBA), 2-carboxyethaneboronic acid,1,2-dicarboxyethaneboronic acid, .beta. beta.′-d carboxyethaneboronate,beta.gamma.-dicarboxypropaneboronate, 2-nitro- and4-nitro-3-succinamidobenzene boronic acids,3-nitro-4-(6-aminohexylamido)phenyl boronic acid,{4-[(hexamethylenetetramine) methyl]phenyllboronic acid,4-(N-methyl)carboxamidobenzene boronic acid,2-{[(4-boronphenyl)methyl]-ethylammonio}ethyl and 2-{[(4-boronphenyl)methyl]diethylammoniol-ethyl groups, succinyl-3-aminophenylboronic add,6-aminocaproyl-3-aminophenylboronic acid, 3-(N succinimidoxycarbonyl)aminophenyl borona-.e, p-(omega.-aminoethyl)phenylboronate, p-vinylbenzeneboronate, N-(3-dihydroxyborylphenyl)succinamic acid,N-(4-rtro-3-dihydroxy borylphenyl)succinamic acid,0-dimethylaminomethylbenzene boronic acid, 4-carboxy benzene boronicacid, 4-(N-octyl)carboxamido benzene boronic acid,3-nitro-4-carboxybenzeneboronic acid, 2-nitro-4-carboxybenzene boronicacid, 4-bromophenylboronate, p-vinylbenzene boronate,4-(.omega.-aminoethyl)phenylboronate, catechol[2-(diethylamino)carbonyl, 4-bromomethyl]phenyl boronate, and5-vinyl-2-dimethylaminomethyl benzeneboronic acid.
 8. The method ofclaim 4, wherein said polylysine is selected from the group consistingof poly-D-lysine (PDL), poly-DL-lysine, poly-e-CBZ-D-lysine,poly-e-CBZ-DL-lysine, and poly-e-CBZ-L-lysine).
 9. The method of claim4, wherein said polyornithine is selected from the group consisting ofpoly-DL-ornithine, poly-L-ornithine or poly-S-CBZ-DL-ornithine.